Autoantibodies directed against intracellular antigens are characteristic features of a number of human diseases. The overall goal of this application is to dissect the autoimmune response to the Golgi complex via the characterization of four unique cDNAs identified recently in the investigator's laboratory. The 4 new Golgi antigens of 95, 97, 160 and 180 kD are referred to as golgin-95, golgin-97, golgin-160, and golgin-180, respectively. These represent 4 of the 5 known Golgi autoantigens cloned to date. The first specific aim has two parts. Part 1: Molecular Analysis of Golgi Autoantigens. The goal is to obtain full-length cDNAs for all 4 golgins. Interestingly, the amino acid sequences deduced for the Golgi autoantigens so far are essentially all coiled-coil domains, which are generally thought to function in protein-protein interaction as in dimer or oligomer formation. Genomic analyses, including mapping of exon-intron junctions and chromosomal loci, will be performed as necessary to completely explain the observed expression of golgins. Part 2: Structural Organization, Interactions, and Immunolocalization of Golgi Autoantigens. 2a. Expression of golgins.Immuno-fluorescence, immunoblotting, Northern blotting and RT-PCR will be used to determine if golgins are differentially expressed in human and mouse tissues and cultured cells. The subcellular location of golgins will be identified by immunoelectron microscopy. The ultrastructural localization of Golgi complex autoantigens will yield information about their structural and functional relationship to each other and to known Golgi components such as beta-COP. 2b. Biochemical and functional analysis of golgins. Physicochemical techniques will be used to determine if golgins form hetero-oligomers or homo-oligomers. Domains responsible for dimer or oligomer formation will be mapped. The protein domains responsible for the localizations or "targeting" to the Golgi complex will be studied by transfection of epitope-tagged golgin cDNA constructs containing complete and truncated fragments. Specific Aim 2 will test the hypothesis that Golgi proteins which are "autoantigens" are components of subcellular macromolecular complexes (particles) which are contributing to common or shared functions. The realization of these aims is enhanced by the availability of the isolated cDNAs and specific antibodies to each of the 4 golgins.